Abstract For decades, the treatment of cancer has relied on surgical resection, chemotherapy and/or radiotherapy. Recently, a number of immune based therapies have provided promising new approaches for cancer treatment. The two most potent immunotherapies are monoclonal antibody (mAB) based bi-specific proteins (eg Bi- specific T cell engagers ? BiTE) and engineered Chimeric Antigen Receptor T cells (CAR T). Both act by inducing T cell mediated killing of cancer cells and have shown remarkable clinical activity, with complete response rates as high as ~90% for B cell malignancies. However, applying these two therapeutic approaches to the vast majority of cancer types is prevented by multiple factors. First, there are only a small number of known cell-surface proteins that are sufficiently specific to cancer to safely allow targeting by antibodies. This is particularly true for solid cancers, where unlike hematopoietic malignancies, loss of healthy cells cannot be readily replenished by stem cell progenitors. Second, as each individual bi-specific protein and/or CAR T cell can only target a single cancer type, different bi-specific and/or CAR T cells will need to be developed for each cancer type. This greatly increases development time and costs. Third, neither therapy is able to effectively target the most abundant and widely expressed cancer antigens known, namely Tumor associated cancer antigens (TACA's). Many cancer specific antigens are not proteins, but rather complex carbohydrates that have limited or no expression in normal tissues. Indeed, altered glycosylation is a near universal feature of cancer. However, generation of monoclonal antibodies specific to complex carbohydrates has proven to be very challenging, greatly limiting their usefulness as targets for cancer immunotherapy. Here we propose to address these issues and develop a novel class of immunotherapeutics that target a carbohydrate antigen common to the vast majority of solid and hematopoietic cancers but not expressed in normal cells. We have termed these molecules as Glycan-dependent T cell Recruiter (GlyTR) technology. Critically, GlyTR technology does not utilize antibodies to target carbohydrate cancer antigens. Preliminary data demonstrates that an O-linked carbohydrate targeted bi-specific GlyTR protein induces T cell dependent killing of human cancer cells in vitro without off-target killing of normal cells. To further develop this technology, we propose the following Aims. Aim 1 optimizes an O-linked glycan targeted GlyTR bi-specific protein for activity and drug development. Aim 2 explores the efficacy and safety of the optimized O-linked glycan targeted GlyTR bi-specific protein. If successful, this work will lead to IND enabling studies of an entire new class of immunotherapeutic cancer killing therapeutics that can target multiple solid and hematopoietic cancers with minimal toxicity.